LOCUS       sequence-269430-        7992 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector for constitutive expression of iRFP670-NLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7992)
  TITLE     Madison Lab plasmids
REFERENCE   2  (bases 1 to 7992)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: BII-ChPtW-iRFP670
FEATURES             Location/Qualifiers
     source          1..7992
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..24,25..31)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    39..55
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     44..66
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     63..79
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     63..79
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   570..604
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(873..889)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(873..889)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             2440..2493
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2500..2526
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             2500..2526
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2533..3477
                     /codon_start=1
                     /product="bright monomeric near-infrared fluorescent 
                     protein engineered from a bacterial phytochrome, with an 
                     emission maximum of 670 nm (Shcherbakova et al., 2016)"
                     /label=miRFP670
                     /label=ORF frame 1
                     /translation="MVAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDH
                     RVIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTE
                     YCGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTG
                     YDRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVT
                     YQPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLV
                     VCHHYLPRFIRFELRAICKRLAERIATRITALES"
     CDS             3481..3501
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3533..4121
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3586..3606)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4004..4015)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4201..4335
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4225..4244)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4279..4298
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5114..5176
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5306..5323)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5306..5322)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5315..5337)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5464..5919
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5551..5570)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5761..5782
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5945..6049
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6050..6118,6119..6910)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6268..6287)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7081..7669
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7570..7589
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7823..7840
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7957..7978
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
       61 cacaggaaac agctatgacc atgattacgc caagcgcgcc cgccgggtaa ctcacggggt
      121 atccatgtcc atttctgcgg catccagcca ggatacccgt cctcgctgac gtaatatccc
      181 agcgccgcac cgctgtcatt aatctgcaca ccggcacggc agttccggct gtcgccggta
      241 ttgttcgggt tgctgatgcg cttcgggctg accatccgga actgtgtccg gaaaagccgc
      301 gacgaactgg tatcccaggt ggcctgaacg aacagttcac cgttaaaggc gtgcatggcc
      361 acaccttccc gaatcatcat ggtaaacgtg cgttttcgct caacgtcaat gcagcagcag
      421 tcatcctcgg caaactcttt ccatgccgct tcaacctcgc gggaaaaggc acgggcttct
      481 tcctccccga tgcccagata gcgccagctt gggcgatgac tgagccggaa aaaagacccg
      541 acgatatgat cctgatgcag ctagattaac cctagaaaga tagtctgcgt aaaattgacg
      601 catgcattct tgaaatattg ctctctcttt ctaaatagcg cgaatccgtc gctgtgcatt
      661 taggacatct cagtcgccgc ttggagctcc cgtgaggcgt gcttgtcaat gcggtaagtg
      721 tcactgattt tgaactataa cgaccgcgtg agtcaaaatg acgcatgatt atcttttacg
      781 tgacttttaa gatttaactc atacgataat tatattgtta tttcatgttc tacttacgtg
      841 ataacttatt atatatatat tttcttgtta tagatccact agttctagag ggacagcccc
      901 cccccaaagc ccccagggat gtaattacgt ccctcccccg ctagggggca gcagcgagcc
      961 gcccggggct ccgctccggt ccggcgctcc ccccgcatcc ccgagccggc agcgtgcggg
     1021 gacagcccgg gcacggggaa ggtggcacgg gatcgctttc ctctgaacgc ttctcgctgc
     1081 tctttgagcc tgcagacacc tggggggata cggggaaaag gcctccaagg cctgcaggag
     1141 tcaatgggaa aaacccattg gagccaagta cactgactca atagggactt tccattgggt
     1201 tttgcccagt acataaggtc aatagggggt gagtcaacag gaaagtccca ttggagccaa
     1261 gtacattgag tcaataggga ctttccaatg ggttttgccc agtacataag gtcaatggga
     1321 ggtaagccaa tgggtttttc ccattactga catgtatact gagtcattag ggactttcca
     1381 atgggttttg cccagtacat aaggtcaata ggggtgaatc aacaggaaag tcccattgga
     1441 gccaagtaca ctgagtcaat agggactttc cattgggttt tgcccagtac aaaaggtcaa
     1501 tagggggtga gtcaatgggt ttttcccatt attggcacat acataaggtc aataggggtg
     1561 actagtggag aagagcatgc ttgagggctg agtgcccctc agtgggcaga gagcacatgg
     1621 cccacagtcc ctgagaagtt ggggggaggg gtgggcaatt gaactggtgc ctagagaagg
     1681 tggggcttgg gtaaactggg aaagtgatgt ggtgtactgg ctccaccttt ttccccaggg
     1741 tgggggagaa ccatatataa gtgcagtagt ctctgtgaac attcggttta gtgaacctgc
     1801 aggtcggccg ccacgaccgg tgccaccatg actgaataca aaccaactgt tcgcctggca
     1861 actcgtgatg atgttccacg tgcagttcgc accctggctg ctgcatttgc tgactaccct
     1921 gcaacccgtc acactgtgga cccagaccgc cacattgaac gtgtgactga actgcaggag
     1981 ctgttcctga cccgtgtggg cctggacatt ggcaaagtgt gggtggcaga tgatggtgct
     2041 gctgtggcag tgtggaccac ccctgaatct gttgaagctg gtgcagtgtt tgctgagatt
     2101 ggcccacgca tggcagaact gtctggcagc cgcctggcag cacaacagca gatggaaggt
     2161 ctgctggcac cacaccgccc aaaagaacct gcttggttcc tggcaactgt gggtgtgagc
     2221 cctgaccacc agggtaaggg cctgggctct gcagtggtgc tgcctggtgt ggaagcagct
     2281 gaacgtgcag gtgtgcctgc tttcctggag acctcagctc cacgcaacct gcctttctat
     2341 gaacgcctgg gcttcactgt gactgctgat gtggaagtgc cagaaggccc acgcacttgg
     2401 tgcatgactc gcaaaccagg tgctagcctt ggctccggcg agggcagggg aagtcttcta
     2461 acatgcgggg acgtggagga aaatcccggc ccactcggct acccctacga cgtgcccgac
     2521 tacgccagcg gcatggtagc aggtcatgcc tctggcagcc ccgcattcgg gaccgcctct
     2581 cattcgaatt gcgaacatga agagatccac ctcgccggct cgatccagcc gcatggcgcg
     2641 cttctggtcg tcagcgaaca tgatcatcgc gtcatccagg ccagcgccaa cgccgcggaa
     2701 tttctgaatc tcggaagcgt actcggcgtt ccgctcgccg agatcgacgg cgatctgttg
     2761 atcaagatcc tgccgcatct cgatcccacc gccgaaggca tgccggtcgc ggtgcgctgc
     2821 cggatcggca atccctctac ggagtactgc ggtctgatgc atcggcctcc ggaaggcggg
     2881 ctgatcatcg aactcgaacg tgccggcccg tcgatcgatc tgtcaggcac gctggcgccg
     2941 gcgctggagc ggatccgcac ggcgggttca ctgcgcgcgc tgtgcgatga caccgtgctg
     3001 ctgtttcagc agtgcaccgg ctacgaccgg gtgatggtgt atcgtttcga tgagcaaggc
     3061 cacggcctgg tattctccga gtgccatgtg cctgggctcg aatcctattt cggcaaccgc
     3121 tatccgtcgt cgactgtccc gcagatggcg cggcagctgt acgtgcggca gcgcgtccgc
     3181 gtgctggtcg acgtcaccta tcagccggtg ccgctggagc cgcggctgtc gccgctgacc
     3241 gggcgcgatc tcgacatgtc gggctgcttc ctgcgctcga tgtcgccgtg ccatctgcag
     3301 ttcctgaagg acatgggcgt gcgcgccacc ctggcggtgt cgctggtggt cggcggcaag
     3361 ctgtggggcc tggttgtctg tcaccattat ctgccgcgct tcatccgttt cgagctgcgg
     3421 gcgatctgca aacggctcgc cgaaaggatc gcgacgcgga tcaccgcgct tgagagcggg
     3481 cccaagaaga agcgcaaggt gggcgcctaa ggctagagtt cgcgtctgga acaatcaacc
     3541 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     3601 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     3661 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     3721 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     3781 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     3841 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     3901 tgacaattcc gtggtgttgt cggggaagct gacgtccttt ccatggctgc tcgcctgtgt
     3961 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     4021 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     4081 ccctcagacg agtcggatct ccctttgggc cgcctccccg cctggaatta attctggcta
     4141 agcacttcgt ggccgtcgat cgtttaaagg gaggtagtga gtcgagatat cgatactagt
     4201 gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     4261 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     4321 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     4381 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtatggc tgattatgat
     4441 cctctagagg gacagccccc ccccaaagcc cccagggatg taattacgtc cctcccccgc
     4501 tagggggcag cagcgagccg cccggggctc cgctccggtc cggcgctccc cccgcatccc
     4561 cgagccggca gcgtgcgggg acagcccggg cacggggaag gtggcacggg atcgctttcc
     4621 tctgaacgct tctcgctgct ctttgagcct gcagacacct ggggggatac ggggaaaaag
     4681 ccataccaat ggagctcacg gggacagccc ccccccaaag cccccaggga tgtaattacg
     4741 tccctccccc gctagggggc agcagcgagc cgcccggggc tccgctccgg tccggcgctc
     4801 cccccgcatc cccgagccgg cagcgtgcgg ggacagcccg ggcacgggga aggtggcacg
     4861 ggatcgcttt cctctgaacg cttctcgctg ctctttgagc ctgcagacac ctggggggat
     4921 acggggaaaa agaattcgat aaaagttttg ttactttata gaagaaattt tgagtttttg
     4981 ttttttttaa taaataaata aacataaata aattgtttgt tgaatttatt attagtatgt
     5041 aagtgtaaat ataataaaac ttaatatcta ttcaaattaa taaataaacc tcgatataca
     5101 gaccgataaa acacatgcgt caattttaca catgattatc tttaacgtac gtcacaatat
     5161 gattatcttt ctagggttaa tctagctgcg tgttctgcag cgtgtcgagc atcttcatct
     5221 gctccatcac gctgtaaaac acatttgcac cgcgagtctg cccgtcctcc acgggttcaa
     5281 aaacgtgaat gaacgaggcg cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa
     5341 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
     5401 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     5461 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     5521 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5581 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5641 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     5701 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5761 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     5821 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     5881 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     5941 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     6001 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     6061 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     6121 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     6181 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     6241 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     6301 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     6361 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     6421 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     6481 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     6541 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     6601 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     6661 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6721 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6781 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6841 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6901 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6961 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     7021 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     7081 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     7141 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     7201 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     7261 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     7321 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     7381 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     7441 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     7501 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     7561 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     7621 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     7681 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     7741 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     7801 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     7861 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     7921 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     7981 ggcaccccag gc
//
